Kees Nooter

Association of enzyme and transporter genotypes with the pharmacokinetics of imatinib

Our objective was to explore the relationships between imatinib pharmacokinetics and 9 allelic variants in 7 genes coding for adenosine triphosphate‐binding cassette transporters (ABCB1 and ABCG2) and enzymes (cytochrome P450 [CYP] 2C9, CYP2C19, CYP2D6, CYP3A4, and CYP3A5) of putative relevance for imatinib.

Molecular profiling of platinum resistant ovarian cancer

The aim of this study is to discover a gene set that can predict resistance to platinum‐based chemotherapy in ovarian cancer. The study was performed on 96 primary ovarian adenocarcinoma specimens from 2 hospitals all treated with platinum‐based chemotherapy. In our search for genes, 24 specimens of the discovery set (5 nonresponders and 19 responders) were profiled in duplicate with 18K cDNA microarrays. Confirmation was done using quantitative RT‐PCR on 72 independent specimens (9 nonresponders and 63 responders). Sixty‐nine genes were differentially expressed between the nonresponders (n = 5) and the responders (n = 19) in the discovery phase. An algorithm was constructed to identify predictive genes in this discovery set. This resulted in 9 genes (FN1, TOP2A, LBR, ASS, COL3A1, STK6, SGPP1, ITGAE, PCNA), which were confirmed with qRT‐PCR. This gene set predicted platinum resistance in an independent validation set of 72 tumours with a sensitivity of 89% (95% CI: 0.68–1.09) and a specificity of 59% (95% CI: 0.47–0.71)(OR = 0.09, p = 0.026). Multivariable analysis including patient and tumour characteristics demonstrated that this set of 9 genes is independent for the prediction of resistance (p < 0.01). The findings of this study are the discovery of a gene signature that classifies the tumours, according to their response, and a 9‐gene set that determines resistance in an independent validation set that outperforms patient and tumour characteristics. A larger independent multicentre study should further confirm whether this 9‐gene set can identify the patients who will not respond to platinum‐based chemotherapy and could benefit from other therapies.

Lack of c-kit exon 11 activating mutations in c-KIT/CD117-positive SCLC tumour specimens.

Previous studies have shown that STI571, a selective tyrosine kinase inhibitor of c-KIT, is highly effective in c-KIT/CD117-positive gastrointestinal stromal tumours (GIST), especially those that have activating mutations in the c-kit exon 11 that encodes the juxtamembrane (JM) domain of the c-KIT oncoprotein. We examined the prevalence of activating exon 11 c-kit mutations in 26 small-cell lung cancer (SCLC) cases in order to explore whether this disease is also a potential target for treatment with STI571. Expression of c-KIT, estimated by immunohistochemistry, was demonstrated in 14 out of 22 SCLC samples (64%); nine samples showed moderate to strong staining (41%), five samples were weakly positive (23%), whereas eight samples (36%) were negative for CD117. Next, we examined the mutational status of exon 11 of the c-kit gene, by single-stranded conformational polymorphism (SSCP) and sequencing in all of the cKIT/CD117-positive tumours. However, no activating mutations in the c-kit exon 11 were found by either technique. Apparently, c-KIT oncoprotein expression in SCLC was not correlated with activating mutations in c-kit exon 11. In analogy to GISTs, our results could imply that SCLC patients would not benefit from treatment with STI571.

No evidence of known types of human papillomavirus in squamous cell cancer of the oesophagus in a low-risk area

Controversial results regarding the presence and role of human papillomavirus in the development of oesophageal squamous cell carcinoma have been published. We used multiple broad-spectrum polymerase chain reactions to identify HPV DNA in oesophageal carcinomas from a low-incidence area. Paraffin embedded- and snap-frozen specimens from oesophageal cancer tissues of 63 patients were examined with a PCR technique with several primer pairs, capable of detecting most known HPV types. In none of the oesophagus cancer tissues could HPV DNA be detected. The role of HPV in this type of carcinoma in a low incidence area remains unclear.

Preclinical evaluation of alternative pharmaceutical delivery vehicles for paclitaxel

New solubilizers, including Sorporol 230, Sorporol 120Ex, Aceporol 345-T, Aceporol 460 and Riciporol 335, as potential new delivery vehicles for paclitaxel were investigated, since recent studies have shown that the paclitaxel delivery vehicle Cremophor EL significantly alters the pharmacokinetics of paclitaxel. Cremophor EL and Tween 80 were used as a reference. As in the case of Cremophor EL, alteration of blood distribution of paclitaxel occurred in the presence of all tested vehicles. Also, no differences in the affinity of paclitaxel for the tested solubilizers was found during equilibrium dialysis experiments. The different vehicles could be distinguished by a different rate of esterase-mediated breakdown, which was correlated with the fatty acid content of the solubilizers. The activation of the complement cascade was less pronounced for all solubilizers, except Riciporol 335, compared to Cremophor EL. The strategies presented here provide the possibility to rapidly screen future candidate delivery vehicles with optimal characteristics for use as a solubilizer in clinical formulations of paclitaxel or other poorly water-soluble drugs.

Differential modulation of cisplatin accumulation in leukocytes and tumor cell lines by the paclitaxel vehicle Cremophor EL

BACKGROUND Several clinical studies have shown that polychemotherapy with the taxanes paclitaxel or docetaxel preceded or followed by cisplatin is associated with important schedule-dependent differences in toxicities, such as leukocytopenia. In general, the pharmacokinetics of both drugs during the combined treatment are unaltered, suggesting that a pharmacodynamic interaction might have occurred. MATERIALS AND METHODS In order to gain insight into this pharmacologic interaction, we performed in vitro drug accumulation studies using peripheral blood leukocytes and a panel of tumor and non-malignant cell lines with paclitaxel and docetaxel, as well as with their respective formulation vehicles Cremophor EL and Tween 80. RESULTS Our results show a significant reduction in the intracellular cisplatin concentration in leukocytes of up to 42% in the presence of Cremophor EL and Tween 80 as compared to the control. This pharmacodynamic interaction of these surfactants with cisplatin seems to be specific for haematopoietic cells, and does not occur in solid tumor cells. CONCLUSION The present data suggest that the pharmaceutical vehicles Cremophor EL and Tween 80 might contribute to the reduced cisplatin-associated myelotoxicity observed in the clinical combination chemotherapy studies with paclitaxel and docetaxel.

Irinotecan pharmacokinetics-pharmacodynamics: the clinical relevance of prolonged exposure to SN-38

We have shown previously that the terminal disposition half-life of SN-38, the active metabolite of irinotecan, is much longer than earlier thought. Currently, it is not known whether this prolonged exposure has any relevance toward SN-38-induced toxicity. Here, we found that SN-38 concentrations present in human plasma for up to 3 weeks after a single irinotecan infusion induce significant cytotoxicity in vitro. Using pharmacokinetic data from 26 patients, with sampling up to 500 h, relationships were evaluated between systemic exposure (AUC) to SN-38 and the per cent decrease in absolute neutrophil count (ANC) at nadir, or by taking the entire time course of ANC into account (AOC). The time course of SN-38 concentrations (AUC500 h) was significantly related to this AOC (P<0.001). Based on these findings, a new limited-sampling model was developed for SN-38 AUC500 h using only two timed samples: AUC500 h=(6.588×C2.5 h)+(146.4×C49.5 h)+15.53, where C2.5 h and C49.5 h are plasma concentrations at 2.5 and 49.5 h after start of infusion, respectively. The use of this limited-sampling model may open up historic databases to retrospectively obtain information about SN-38-induced toxicity in patients treated with irinotecan.

Modulation of cisplatin pharmacodynamics by Cremophor EL: experimental and clinical studies

The paclitaxel vehicle Cremophor EL (CrEL) has been shown to selectively inhibit the accumulation of cisplatin in peripheral blood leucocytes, but not in tumour cells in vitro, and we hypothesised that this phenomenon is responsible for the improvement of the therapeutic index of cisplatin observed in combination studies with paclitaxel. Here, we report on studies assessing the interaction between CrEL and cisplatin in a murine model, and involving the potential clinical applicability of CrEL as a protector for cisplatin-associated haematological side-effects. In mice, CrEL (0.17 ml/kg, intravenous (i.v.)) given in combination with cisplatin (10 mg/kg, intraperitoneal (i.p.)) did not change the pharmacokinetics of cisplatin. Cisplatin-induced haematological toxicity, expressed as white blood cells (WBC) at nadir, was significantly reduced by CrEL from 5.05+/-0.95 to 6.50+/-1.31 x 10(9)/l (P=0.0009). Data obtained from cancer patients treated with cisplatin (70 mg/m(2), 3-h i.v.) and topotecan (0.45 or 0.60 mg/m(2)/day x 2) preceded by CrEL (12 ml, 3-h i.v.) (n=6) or without CrEL (n=10) similarly indicated significant differences in the percent decrease in WBC between the groups (46.5+/-18.7 versus 67.2+/-15.0%; P=0.029). Likewise, the percent decrease in platelet count was significantly greater in the absence of CrEL (23.9+/-5.38 versus 73.3+/-15.5%; P=0.0003). Pharmacokinetic parameters of unbound and total cisplatin and of topotecan lactone and total drug were not significantly different from historic control values (P>or=0.245). Overall, this study provides further evidence on the important role of CrEL in the pharmacological and toxicological profile of cisplatin, and implies that reformulation of cisplatin with CrEL for systemic treatment might achieve an improvement of its therapeutic index, particularly in the setting of a weekly dose-dense concept.

Detection of human C-type "helper" viruses in human leukemic bone marrow with murine sarcoma virus-transformed human and rat non-producer cells.

Bone‐marrow cells from two leukemic children were co‐cultivated with the canine thymus cell line A 7573. In early passages, C‐type oncornaviruses were released as detected by extracellular reverse transcriptase assay. Co‐cultivation of the infected canine cells with the non‐producing cell lines R‐970‐5 (human) or K‐NRK (rat) both transformed by Kirsten mouse sarcoma virus (MSV) yielded a new pseudotype of MSV that could transform rat embryo, rabbit SIRC and human kidney cells but not mouse embryo cells. The focus formation could be inhibited by an antiserum to the simian sarcoma virus but not by a serum directed against murine leukemia virus. A cell line derived from a focus of transformed cells became a high virus producer. By immunofluorescence it could be established that the virus is related to the simian sarcoma virus. It is concluded that the leukemic bone‐marrow cells produce a C‐type oncornavirus that can serve as a helper virus to the defective MSV.

Transfection of mouse bone marrow cells with Rauscher virus proviral DNA

Abstract Mouse bone marrow cells were transfected with DNA isolated from Rauscher virus-induced leukemic spleen cells. Successful transfection was manifested by virus production and transformation. Virus production by the transfected bone marrow cells was shown by the XC test. Injection of transfected cells into syngeneic hosts led to the development of erythrobastosis, probably due to the spread of virus reproduced by infectious proviral DNA. The transforming potential of the infectious viral DNA was demonstrated by the formation of erythroid colonies in vitro without the addition of erythropoietin to the culture medium.